SUMMARY Extracellular and intracellular pathogen recognition by members of the TLR and NLR families initiate innate immune and inflammatory responses. TLRs and NLRs both activate the NF-kB and MAPK signaling pathways leading to the transcription and secretion of multiple inflammatory cytokines (e.g. TNF, IL-6, and IL-8). Unlike TLRs, NLRs can also activate multiprotein inflammasome complexes that processes proIL-1 and proIL-18 to their active forms. In less than five years, NLRs have been linked to hereditary autoinflammatory diseases and innate immune response towards a growing number of pathogens, including Bacillus anthracis, Yersinia pestis, and Francisella tularensis. Inflammasome assembly results from interaction of a pyrin containing NLR protein with ASC (apoptotic speck protein with a CARD) followed by recruitment and activation of Caspase-1. The inflammasome complex may also be involved in inducing a form of caspase-1 dependent apoptosis. ASC interactions with NLRs and the protein Pyrin can also activate NF-kB. Recently, the discovery of pyrin only proteins (POPs) that influence both NF-kB and inflammasome functions has suggested an avenue for host regulation of the proinflammatory response and pathogen subversion thereof. The long-term objective of this proposal is to understand the molecular regulation of innate immune responses dependent upon pyrin only proteins. Further, understanding of inflammasome regulation will be extended to host-pathogen interactions where inflammasome function or subversion is demonstrated to be critical for immunity or disease. Specifically, we have identified a second human POP (POP2) which inhibits NF-kB p65 activity, interacts with ASC, and prevents the activation of a number of NLR activated inflammasomes. We will test the hypothesis that POP2 negatively regulates proinflammatory responses by interfering with both inflammasome- mediated caspase-1 activation and attenuating NF-kB signals, thus dampening harmful inflammation and preventing or reducing macrophage death during innate immune responses. Combining broad based molecular and in vivo approaches, we will address the following aims: 1) Establish the molecular basis for NF-kB inhibition by POP2 2) Establish the molecular basis for POP2 regulation of inflammasome activation. 3) Establish the in vivo role of POP2 in modulating inflammatory innate immune responses.